Development and verification of a fluorescence quantitative detection method for SARS-CoV-2 antibodies based on magnetic bead sorting and peptide self-assembly
Objective To develop and verify a fluorescence quantitative detection method for antibody against severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)based on magnetic bead sorting and peptide self-assembly.Methods Magnetic beads were specifically bound to nucleocapsid(N)protein of SARS-CoV-2 by click chemistry reaction,which were detected by Western blot and BCA.Using magnetic beads labeled with N protein as coating antigen and donkey anti-human IgG labeled with alkaline phosphatase(ALP)as enzyme-labeled antibody,combined with assemblable peptide and thioflavine-T(ThT)luminescence system,a quantitative detection method for SARS-CoV-2 antibody was established,and the specificity,linear range,limit of detection(LOD),precision and durability were verified.The developed method and commercial ELISA kit were used to detect 8 serum samples with known background respectively,and the consistency between them was calculated.Results Most of SARS-CoV-2 N proteins were stably bound to magnetic beads.The developed method specifically detected positive serum samples.The concentration of SARS-CoV-2 IgG antibody standard ranged from 2.5 to 80 ng/mL,showing a good linear relationship with fluorescence value,and the linear equation was y=182.11 x+5 095.5,R2=0.99;The LOD was 2.5 ng/mL;The coefficient of variation(CV)within batches ranged from 3.9％to 10.6％,the CV between batches ranged from 6.0％to 13.8％,and the CV of durability was within 1.3％～6.2％.The consistency between the results of the developed method and commercial ELISA kit was high(R2=0.99,P＜0.000 1).Conclusion The developed fluorescence quantitative detection method based on magnetic bead sorting and peptide self-assembly showed good specificity,precision and durability,which might be used for quantitative detection of SARS-CoV-2 antibody.
Severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)AntibodyMagnetic bead sortingSelf-assem-bly of peptides
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